How regeneration occurs is a fundamental problem in biology. One of the earliest known steps in response to wounding is stem cell proliferation. Proliferation of stem cells is required for regeneration of any missing or damaged tissues. However, how missing tissue is detected remains unknown. Planarians are fresh-water flatworms renowned for their remarkable ability to regenerate, restoring missing tissue efficiently even after massive trauma. A near-finished genome sequence of the planarian S. mediterranea and the recent development of molecular tools for this model system have enabled new research into its regenerative powers. Whereas all wounds stimulate stem cell proliferation throughout the body of planarians, only wounds involving missing internal tissues stimulate stem cell proliferation near the wound. I propose to exploit this differential stem cell behavior to uncover the molecular mechanisms planarians use to detect missing tissue. Specifically, I will 1) identify genes that are required for signaling missing tissue by performing microarrays involving a specific set of wound types. The wound set was selected to challenge the animals from distinct starting contexts and to allow comparisons of gene expression changes between wounds with and without missing tissue; 2) determine functional roles of these genes in response to missing tissue using RNAi-based screens combined with immunohistochemical staining; and 3) establish the sequence of molecular events following removal of tissue during wounding, using a combination of expression analyses, RNAi, and cell biological approaches. Given the high level of gene conservation between planaria and humans, elucidation of factors that mediate communication between wounds and stem cells will broadly enhance our understanding of stem cell biology and may provide potential therapeutic targets for regeneration. Stem cell proliferation in the absence of missing or damaged tissue can lead to cancer, and stem cells have now been implicated in re-growth of several types of tumors after treatment. Thus, understanding the signals that govern stem cell proliferation may also provide new insights into cancer mechanisms.